The goal of the proposed project is to study protein fiber formation using an integrated microfluidic device, and to develop "green" processes for generating performance protein fibers for biomedical applications. Despite significant advances in protein synthesis and fabrication technology, spiders are still the best engineers producing extraordinarily strong silk fibers at low pressures, ambient temperatures, and with water as solvent. It has been postulated that spiders finely control the types of silk proteins, the physiological conditions of protein solutions, as well as the mechanical deformation of the silk thread in order to spin silk fibers of exceptional strength. Here, we hypothesize that the use of microsystem technology provides an important paradigm for studying the mechanisms involved in silk fiber formation and, subsequently, developing "green" processes for producing performance protein fibers. Specifically, recombinant silk-elastin-like proteins (SELPs) comprised of polypeptide sequences derived from silk and elastin will be used as a model material for the proposed project. Specific Aim 1. Determine the protein solution characteristics that influence the fiber formation and the fiber secondary structures. An integrated microfluidic system consisting of PDMS-based microchannels and an aluminum micro heater, for local temperature control, will be fabricated for studying SELP fiber formation. SELP aqueous solution will be pumped into the microfluidic system through the main channel, while salt and acidic or basic solutions will be driven through side channels to adjust the ionic strength and pH value of the SELP solution in a controlled manner. The effects of pH, ions and temperature on the fiber formation and the fiber secondary structures will be monitored using an optical microscope and characterized using a variety of spectroscopic methodologies. Specific Aim 2. Investigate the flow properties of the spinning duct that determine the fiber formation, secondary structures, and mechanical properties of SELPs. Mechanical deformation of the SELP thread will be controlled by narrowing the channel width and varying the flow rates. The resulting mechanical properties and secondary structures of SELP fibers will be characterized. In this effort, computational fluid dynamics (CFD) simulations and a constitutive analytical model will be utilized to enhance our understanding of the effects of shear/elongational flows on fiber formation, structure, and properties. Specific Aim 3. Explore the effects of the protein primary structures in modulating the formation, structure, and properties of SELP fibers. SELPs composed of varying lengths of silk-/elastin-like blocks and crosslinking sites will be synthesized, and their capability to form fibers will be examined. The influence of incorporating lysine residues, the size of silk-like blocks, as well as the ratio of silk- to elastin- like blocks on the formation, structure, and properties of SELP fibers will be investigated. PUBLIC HEALTH RELEVANCE: Microsystem-Based Formation of Recombinant Protein Fibers Spiders have amazed scientists by producing extraordinarily strong silk fibers using "green" processes at ambient temperatures, low pressures and with water as solvent. We propose to use an integrated microfluidic device as an important tool for studying the formation mechanisms of silk fibers and for developing "green" fiber fabrication techniques. Significantly, the demystification of spiders'remarkable silk spinning process and the development of "green" fabrication techniques may lead to the engineering of performance protein fibers for many biomedical applications.